The major hypothesis of this IPCAVD Program is that combinations of mucosal and parenteral immunizations with novel DNA and Sindbis virus delivery systems will induce both cell-mediated and humoral immunity to HIV. Such immune responses may prevent transmissions at the sites of virus entry, decrease systemic viral spread, or alter the disease course following infection. The main objectives of Project 1 are the following. First, to optimize the DNA constructs, expressing SIV and HIV antigens, that are needed for this Program. The optimized DNA, encoding viral antigens, will be used for this Program. The optimized DNA, encoding viral antigens, will be used for both the PLG-DNA vaccines, as well as the Sindbis virus replicons to be used throughout the rest of the Program. Second, to conduct studies in mice that will define the optimal induction of mucosal and systemic immune responses. Third, to investigate the mechanisms of immune induction at mucosal surfaces when using novel mucosal vaccination strategies. Fourth, to study the trafficking mechanisms involved in the induction of HIV-specific local and peripheral immunological responses. And lastly, to study the memory induced in both the humoral and cellular immune response using our novel mucosal vaccination strategies. Importantly, the mucosal immunization studies in mice will be critical for testing large numbers of vaccine variables, so that a tractable number of vaccine protocols can then be evaluated in macaques. The mouse studies will also validate novel expression systems in an in vivo setting in a cost-effective fashion and will allow analysis of mechanisms of immune induction, lymphocyte trafficking and maintenance of immunological memory. Because this Project is part of the larger Program format of the IPCAVD, novel findings in induction of mucosal immune responses in macaques may define novel issues for mechanistic studies in basic immunology in mice. Thus, such a "comparative immunology" approach will be essential for defining both unique as well as shared mucosal immunity issues in both rodents and primates.